Neoplastic Diseases
The transformation of a normal cell into a malignant cell results, among other things, in the uncontrolled proliferation of the progeny cells, which exhibit immature, undifferentiated morphology, exaggerated survival and proangiogenic properties. It is also often associated with overexpression or constitutive activation of oncogenes not normally expressed by normal mature cells. Once a tumor has formed, cancer cells can leave the original tumor site and migrate to other parts of the body via the bloodstream and/or the lymphatic system by a process called metastasis. In this way, the disease may spread from one organ or part to another non-contiguous organ or part.
The increased number of cancer cases reported around the world is a major concern. Currently there are only a handful of treatments available for specific types of cancer and these treatments provide only limited efficacy. In order to be most effective, these treatments require: early detection of the malignancy, reliable assessment of its severity, methods of tracking possible metastasis, and readouts to monitor patients' response to treatment.
The Hedgehog Pathway
The seven transmembrane domain containing protein smoothened (SMO) serves as the key player for signal transduction of the Hedgehog (Hh) pathway. There are 3 Hh-related genes in vertebrates: sonic hedgehog (Shh), Indian Hedgehog and Desert Hedgehog. Hh is used here to encompass all three genes. However, the pathway's function is inhibited by a Hh-binding twelve transmembrane domain protein, patched (PTC), in the absence of Hh ligands. There are two homologs of patched in vertebrates (e.g., Ptch1 and Ptch2 in mouse, PTCH1 and PTCH2 in human), and one gene in Drosophila. As used herein, “Ptc” is used to designate the human gene or the mouse gene. The context specifies which species was used. It has recently been shown that the type I transmembrane proteins CDON (cell-adhesion-molecule-related/downregulated by oncogenes) and BOC (biregional CDON-binding protein; also known as “brother of CDON”) are also able to bind Hh (Okada, A et al., Nature 444 (7117): 369-373). In the presence of active Hh ligands, it is thought that binding of Hh to PTC and BOC/CDON complexes releases their inhibition of SMO, allowing SMO to signal downstream to Gli/Ci transcription factors. As transcription factors, Gli molecules can regulate the expression of Hh target genes by directly associating with a specific consensus sequence located in the promoter region of the target genes such as GLI1 itself, PTCH1 and HIP (McMahon, A P et al., Curr Top Dev Biol 2003, 53: 1-114). FIG. 1 shows a simplified diagram of Hh signaling in the presence of Hh. Hh proteins (sonic hedgehog (Shh), Indian hedgehog (Ihh) (not shown) and desert hedgehog (Dhh) (not shown)) are secreted molecules, functioning both on nearby and distant cells in developing tissues. Following translation, Hh proteins enter the secretory pathway and undergo autoprocessing and other post-translational modifications. Binding of Hh to PTC1 alone, in the absence of interaction with BOC/CDON complexes, results in lower levels of hedgehog pathway activation. Interaction of Hh with both PTC and BOC/CDON complexes results in increased hedgehog pathway activation. Other major components includes Su(Fu), REN, Costal2, Fused and recently Gas1. The C-terminal tails of BOC/CDON might convey other signals to cytoskeletal or integral membrane proteins that induce morphological changes in the cell and signal to the nucleus through unknown mechanisms.
CDON and BOC are type I transmembrane receptors consisting of four or five immunoglobulin (Ig) and two or three fibronectin type III (FNIII) repeats in the extracellular domain, and an intracellular domain with no identifiable motifs. This domain architecture is closely related to that of axon guidance receptors of the Robo and DCC (deleted in colorectal cancer) families (FIG. 2). Both CDON and BOC share a high degree of homology in their extracellular domains and are expressed during early stages of development of the central nervous system (Okada et al., 2006. Nature, 444: 369-373). CDON and BOC form complexes with each other in a cis fashion.
Cerebellum Development and Medulloblastoma
During cerebellum development, Granule Cell Precursors (GCP) undergo rapid proliferation in the External Germinal Layer (EGL) of the cerebellum. Expansion of the GCP is dependent of Sonic hedgehog secreted by Purkinje cells. After their final division, GCP differentiate into Granule cells (GC) and migrate through the Purkinje cell layer to form mature granule cells that reside in the internal granule cell layer (IGL).
Medulloblastoma is an embryonal neuroepithelial tumor of the cerebellum and the most common malignant brain tumor occurring in children (Reviewed in Rossi et al. 2008. Clin. Cancer. Res, 14: 971-976). The tumors develop in the cerebellum, in a part of the brain called the posterior fossa, but may spread to other parts of the brain including the spinal cord. Medulloblastomas may also spread to other parts of the body often through the cerebrospinal fluid (CSF), which surrounds and protects the brain and spinal cord.
It is common knowledge that a clinical diagnosis of medulloblastoma is based on a combination of symptoms and signs observed in the evolution of the disease in the patient. An earlier diagnosis, particularly during neurosurgical clinical examination, would greatly increase the chances of successful treatment and recovery. Currently, proper diagnosis of medulloblastoma involves an invasive procedure whereby a biopsy is performed and sample cells are removed from the tumour and subsequently analyzed. In addition, CSF can be examined for the presence of tumor cells to determine if the tumor has metastasized, which is usually a very poor prognostic (Reviewed in Rossi et al. 2008, supra).
Despite improved multimodal treatment regimens, approximately one-third of patients with medulloblastoma remain incurable and current treatments significantly damage long-term survivors. Although fewer than 500 children per year in the US are diagnosed with medulloblastoma, the outcome is almost invariably poor. Surgery with subsequent radiation or chemotherapy has increased survival to greater than 50%, but the current methods of treatment result in severe long-term side effects including mental retardation.
Medulloblastoma can be a very aggressive tumor. The treatment of Medulloblastoma consists of surgery to remove as much tumor mass as possible, followed by radiotherapy, sometimes also combined with chemotherapy. In spite of these very aggressive treatments, only about 50 to 70% of patients survive after 5 years. For the survivors, the intense radiotherapy often leads to severe side effects, such as permanent cognitive impairment. However, not all MB are equally aggressive and information which informs prognosis, such as risk-assessment by the presence or absence of a molecular marker, would be useful to help determine which type of treatment (less or more aggressive) is most appropriate for each patient.
After surgery and radiotherapy, Medulloblastoma patients are closely followed by MRI for potential relapse of the tumor. This is particularly important in patients in which total resection of the tumor was impossible, which constitute 30-40% of the total number of patients undergoing surgery. Here, having a rapid, simple and robust method to detect tumor relapse at early stages would be very useful.
In some cases, tumor cells might disseminate in the CSF and metastasize to the spinal cord and other parts of the brain. This is one of the most feared complications of MB. The sensitive detection of MB disseminating cells in the CSF or the detection of metastases at early stage would certainly improve treatment outcome.
To improve the outcome of those medulloblastoma patients with high-risk disease as well as the quality of life of all survivors following treatment, both novel therapies and improved tumor classification are required. Novel therapies will result from a greater understanding of the disease process and are likely to involve small molecules designed to target specific pathways, including hedgehog, that become dysregulated during oncogenesis. An improved tumor classification will incorporate an assessment of the molecular profiles of medulloblastomas with defined biological behaviors or of the status of cellular pathways that are potential targets for novel therapies.
Thus, there is a need for the identification of novel targets that could serve as biomarkers of early stages of cancer. There is a need for the identification of novel biomarkers for better characterization and classification of tumors. Also, there is a need for improved diagnostic and prognostic detection methods as well as novel anti-cancer treatments which can be administered to subjects either suffering from or who are at a high risk of developing cancer to prevent, inhibit or treat the disease and the spread thereof.
The present description refers to a number of documents, the content of which is herein incorporated by reference in their entirety.